Control of gene expression is mediated partly by recruitment of transcriptional factors to the promoter elements. Recruitment of these factors is further regulated by the chromatin structure surrounding the promoter region. If the chromatin structure is in an "open" conformation, the promoter element is accessible to the transcriptional factors but if it is in a "closed" conformation, the promoter element is inaccessible. Using a novel yeast strategy, we cloned a human serine-threonine kinase that appears to convert chromatin from a "closed" conformation to an "open" conformation. This kinase is expressed in all tissue types and is localized to the nucleus of human cells. The mechanism by which this kinase "opens" up the chromatin is currently being investigated. Once the chromatin is opened, transcriptional factors are recruited to activate or suppress transcription. Although a great deal is known about the transcriptional factors, study of their recruitment would be greatly enhanced by a methodology that allows detection of their recruitment in vivo, a methodology that does not exist yet. Toward this end, we developed a novel methodology PIN*POINT (protein position identification with nuclease tail). In this method, a fusion protein composed of a chosen protein linked to a non-sequence-specific nuclease is expressed in vivo and the binding of the protein to DNA is made detectable by the nuclease-induced cleavage near the binding site. Using PIN*POINT, we have been able to demonstrate that the beta-globin LCR (Locus Control Region) promotes recruitment of transcriptional activator Sp1 to the beta-globin promoter in MEL (murine erythroleukemia) cells. In collaboration with Dr. Myung Kim, NIDDK, we are also studying the role of the thyroid hormone receptor and thyroid hormone in recruitment of the general transcription factors.